1. Field of the Invention
This invention relates to novel inhibitors of serine/threonine protein kinases (e.g., AKT and related kinases), pharmaceutical compositions containing the inhibitors, and methods for preparing these inhibitors. The inhibitors are useful for the treatment of hyperproliferative diseases, such as cancer and inflammation, in mammals and especially in humans.
2. Description of the State of the Art
Protein kinases are a class of enzymes that catalyze the transfer of the γ-phosphorate group from ATP to a recipient protein, acting as a substrate. The specific target of the kinase is the hydroxyl group of a serine, threonine or tyrosine residue. As a result of this specific targeting, kinases are generally referred to as serine/threonine protein kinases or tyrosine protein kinases. The human genome is estimated to encode in excess of 500 distinct protein kinases.
The seemingly insignificant task of phosphorylation of a serine, threonine or tyrosine residue belies the importance of protein kinases in the processes of signal transduction and regulation of cellular functions. Kinases are typically mediated by transmembrane cellular receptors, such as G-protein coupled receptors or growth factor receptors, which when activated by extracellular ligands cause the phosphorylation of intracellular proteins. Often, an interconnected series (or cascade) of protein kinases is necessary to exert the overall effect of this initial signal, which can ultimately result in effects as extreme as cell death (apoptosis).
The ratio of phosphorylated to unphosphorylated protein is a delicate equilibrium, with protein phosphatases acting as the negative regulator of protein kinases, removing the phosphoryl group as it is no longer required. As an example of this interplay, the phosphorylation state of kinases can control whether a cell undergoes division, arrests in the cell cycle or programmed cell death. Should this kinase/phosphatase relationship become disregulated, the potential consequences relating to disease are enormous. For example, abnormal protein kinase activity or expression may be correlated with numerous hyperproliferative diseases, inflammation and tissue repair, and has been associated with a large number of diseases ranging from the relatively non-life threatening, such as psoriasis, to those which are almost always fatal, such as glioblastoma multiforme, an aggressive brain cancer.
Significantly, atypical protein phosphorylation and/or expression is often reported to be one of the causative effects of abnormal cellular proliferation, metastasis and cell survival in cancer. The abnormal regulation and/or expression of various kinases, including VEGF, ILK, AKT, ROCK, p70S6K, Bcl, PKA, PKC, Raf, Src, PDK1, ErbB2, MEK, IKK, Cdk, EGFR, BAD, CHK1, CHK2 and GSK3 amongst numerous others, has been specifically implicated in cancer.
Recent data from the CDC indicate that cancer is the second most common cause of death in the United States, with nearly a quarter of all deaths reported being attributable to malignant neoplasms (Anderson, National Vital Statistics Report, 2001, 49 (11):1). Despite recent advances in the understanding of the genesis, progression and treatment of cancer, much still needs to be done to improve the overall prognosis of cancer patients.
The phosphatidylinositol 3′-OH kinase (PI3K) pathway is one of the signaling pathways that exerts its effect on numerous cellular functions including cell cycle progression, proliferation, motility, metabolism and survival. Activation of receptor protein tyrosine kinases (RTKs) cause PI3K to phosphorylate phosphatidylinositol (4,5)-diphosphate[PtdIns (4,5)P2], generating the membrane-bound phosphatidylinositol (3,4,5)-triphosphate[PtdIns(3,4,5)P3]. This in turn promotes the recruitment of a variety of protein kinases from the cytoplasm to the plasma membrane through the binding of PtdIns (3,4,5)P3 to the pleckstrin-homology (PH) domain of the kinase. Kinases notable as key downstream targets of PI3K include phosphoinositide-dependant kinase 1 (PDK1) and AKT (also known as Protein Kinase B.) Phosphorylation of such kinases then permits the activation or deactivation of numerous other pathways involving mediators such as GSK3, mTOR, PRAS40, FKHD, NF-κB, BAD, Caspase-9, etc.
An important negative feedback mechanism for the PI3K pathway is PTEN, a phosphatase that catalyses the dephosphorylation of PtdIns (3,4,5)P3 to PtdIns (4,5)P2 (Furnari, F. B., et al, Cancer Res. 1998, 58:5002; Dahia, P. L. M., Hum. Molec. Genet. 1999, 8:185). It is of enormous significance that in greater than 60% of all solid tumors, PTEN is mutated into an inactive form, permitting the constitutive-activation of the PI3K pathway. As the majority of cancers are solid tumors, such an observation would suggest that by specifically targeting either PI3K itself or the individual downstream kinases in the PI3K pathway, one might able to mitigate the effects of various cancers and restore normal cellular function.
One of the best-characterized targets of the PI3K lipid products is the AGC serine/threonine protein kinase AKT (Hemmings, B. A., Science, 1997, 275:628). AKT is the human homologue of the protooncogene v-akt of the acutely transforming retrovirus AKT8. Its high sequence homology to protein kinases A and C has also earned it the names Protein Kinase B (PKB) and Related to A and C (RAC.) Three isoforms of AKT are known to exist, namely Akt1, Akt2 and Akt3, which exhibit an overall homology of 80% (Staal, S. P, Proc. Natl. Acad. Sci., 1987, 84:5034; Nakatani, K, Biochem. Biophys. Res. Commun., 1999, 257:906). In addition, both Akt2 and Akt3 exhibit splice variants.
Upon recruitment to the cell membrane by PtdInd (3,4,5)P3, AKT is phosphorylated (activated) by PDK1 at T308, T309 and T305 for isoforms Akt1, 2 and 3, respectively, and at S473, S474 and S472 for isoforms Akt1, 2 and 3, respectively. Such phosphorylation occurs by an as yet unknown kinase (putatively named PDK2), although PDK1 (Balendran, A., Curr. Biol., 1999, 9:393), autophosphorylation (Toker, A., J. Biol. Chem., 2000, 275:8271) and integrin-linked kinase (ILK) (Delcommenne, M., Proc. Natl. Acad. Sci. USA, 1998, 95:11211) have been implicated in this process. Although monophosphorylation of AKT activates the kinase, bis(phosphorylation) is required for maximal kinase activity.
AKT is believed to assert its effect on cancer by suppressing apoptosis and enhancing both angiogenesis and proliferation. In addition, AKT has been shown to be overexpressed in many forms of human cancer including, but not limited to, colon (Zinda, et al, Clin. Cancer Res., 2001, 7:2475), ovarian (Cheng, J. Q., et al., Proc. Natl. Acad. Sci. USA, 1992, 89:9267), brain (Haas Kogan, D., et al, Curr. Biol., 1998, 8:1195), lung (Brognard, J., et al, Cancer Res., 2001, 61:3986), pancreatic (Cheng, J. Q., et al., Proc. Natl. Acad. Sci., 1996, 93:3636), prostate (Graff, J. R., et al, J. Biol. Chem., 2000, 275:24500) and gastric carcinomas (Staal, S. P., et al., Proc. Natl. Acad. Sci. USA, 1987, 84:5034).
The development of kinase inhibitors that target abnormally regulated pathways and ultimately result in disease is of enormous ethical and commercial interest to the medical and pharmaceutical community. As such, a compound that inhibits (1) recruitment of AKT to the cell membrane, (2) activation by PDK1 or PDK2, (3) substrate phosphorylation, or (4) one of the downstream targets of AKT would therefore be a valid target as an anticancer agent, either as a stand-alone therapy or in conjunction with other accepted procedures.